: Hepatitis B virus (HBV) causes chronic hepatitis, cirrhosis, and hepatocellular carcinoma (HCC), and is especially common among minorities such as African Americans and Asian Americans. The mechanism by which HBV causes HCC is unclear, but it is likely that HCC results from both specific viral factors and non-specific mutagenesis due to constant cell turnover. One candidate viral factor is the large surface protein, since expression of this protein in transgenic mice results in HCC. We have recently shown that large surface protein can activate cellular genes at the transcriptional level, apparently by forming non-secretable particles in the endoplasmic reticulum (ER) and activating ER stress. Further preliminary data indicate that large surface protein causes cell death, probably by apoptosis. Since hepatocytes containing large surface protein particles within the ER ("ground glass cells") are seen in the livers of people with chronic hepatitis B, our results point to a specific mechanism whereby HBV may cause HCC. However, the reason for ground glass cell formation in infected livers is still unclear. For the next cycle of this project, we propose to study events both up-stream and down-stream of the formation of ground glass cells. First, we wish to validate our hypothesis that mutations in the viral genome can lead to ground glass cells, and determine if two common naturally occurring viral mutants can cause ground glass cells and HCC in transgenic mice. Second, we will clone out HBV genomes from human ground glass cells, and look for additional mutations that may be responsible for ground glass cell formation. Third, we will determine how large surface protein causes cell death, and whether it is by apoptosis. We will also determine if there is increased apoptosis of ground glass cells in transgenic mice, and look for activation of apoptotic pathways in human livers with ground glass cells. These experiments should give novel insights into the mechanism by which HBV causes liver disease.